Transport across the nuclear membrane is necessary at several steps in the life cycle of HIV. Once in the cytoplasm, the viral RNA is converted to double stranded DNA which must enter the nucleus. Viral regulatory proteins enter the nucleus while viral transcripts are exported into the cytoplasm. The viral REV protein has been identified as a key regulator of transport of HIV envelope mRNA. Several approaches have been taken to understand how REV may function. Cell lines are being prepared which express a REV/glucocorticoid receptor chimera which enters the nucleus in an inducible fashion; these cell lines also contain a construct containing a REV response element controlling expression of a chloramphenicolacetyltransferase (CAT) gene. This will allow CAT expression to be conveniently followed allowing a direct measure of export of CAT mRNA into the cytoplasm. A means has also been devised for generating nuclei in vitro around exogenously added DNA. The method uses extracts from Xenopus laevis eggs. The nuclei assembled in such extracts mimic interphase nuclei in many ways and carry out active nuclear transport. These preparations allow examination of the mechanism of REV action and the movement of other molecules involved in the HIV life cycle. In other studies, the structure of the nuclear pore has been examined. The nuclear pore requires glycoprotein components for proper morphology and function. Previous studies indicate the overexpression of nuclear pore glycoproteins may be toxic to cells. To circumvent these problems, the cDNA encoding the major nuclear glycoprotein p62 has been placed under the control of a glucocorticoid responsive expression vector. By expressing the sense and antisense constructs, it should be possible to study the effects of suppression or overexpression of this nuclear pore component. A number of the components of the pore complex have now been molecularly cloned. Understanding how retroviral products cross the nuclear envelope is critical to attempts to regulate or inhibit the critical steps in the HIV life cycle.